DESCRIPTION (appended verbatim from investigator's abstract): Programmed cell death (PCD) plays an indispensable role in the development and maintenance of homeostasis within all multicellular organisms. Thus, the defects in PCD pathways contribute to a variety of diseases including AIDS and oncogenesis. The evolutionarily conserved BCL2 family of proteins are thought to be central regulators, acting at or near a point in the PCD pathway that dictates whether or not cells are committed to die. Members of this family can roughly be divided into two groups; those that promote (BAX) and those that inhibit (BCLXL) apoptosis. Much recent evidence supports the view that critical life/death decisions are regulated by the interaction of BCL2 family members with mitochondria, yet a detailed understanding of many of the important underlying mechanisms and critical biochemical interactions is lacking. Our goal is to use the genetic and biochemical tools available in yeast to identify the mechanisms and interactions that underlie the function of BCL2 family members in caspase independent cell death pathways. All available evidence strongly indicates that BCL2 family members act directly upon highly conserved mitochondrial components in yeast that correspond exactly to their apoptotic substrates in mammalian cells. Our preliminary results are consistent with a model in which BCL2 proteins generate their effects through interaction with a conserved mitochondrial site. Although these interactions have been uncovered in yeast, our underlying general hypothesis is that they are reflective of a similar general relationship in mammalian cells. These results lead to the following specific aims: 1. Test the hypothesis that "BH3 only" proteins function by modifying the interaction of antiapoptotic members with a critical site in mitochondrial membranes. "BH3 only" members of the BCL2 family have all the characteristics expected of proteins which, when translocated to mitochondrial membranes in response to a death signal, function to restrict the ability of BCLXL to associate with critical sites. We propose to directly test this hypothesis. 2. Identify proteins forming the site recognized by BCL2 family members in mitochondria. It is our goal to use genetic approaches to specifically identify proteins forming the "receptor" for BCL2 family members in mitochondria, as well as proteins that function to promote BCLXL mediated cell survival. 3. Determine the mechanisms underlying the ability of BAX to mediate growth arrest in the absence of cell killing. A large body of evidence exists to indicate that cell cycle and cell death processes are interconnected, although the mechanisms and signals that integrate these two processes are completely unknown. Our preliminary studies have allowed us to genetically separate a BAX mediated cell killing process from a growth arrest process. We propose to use genetic strategies to identify the molecular components required for BAX mediated growth arrest, and subsequently, use these mutants to address the question of whether BAX generated cell killing requires growth arrest.